Immunological memory is a defining feature of the adaptive immune system. Memory T cells carry an Imprint from the previous antigen encounter for decades, which enables them to respond more vigorously and to develop effector functions more rapidly upon antigen recognition than naive T cells. Epigenetic mechanisms have been suspected to allow for more rapid gene activation and also maintain a response pattern that was originally developed on the first antigen encounter. With Increasing age the ability of the adaptive immune system to generate and maintain T cell memory is severely impaired compromising the efficacy of vaccination. In studies during the last funding period, we have identified negative feedback signaling loops that impair T cell responses in the elderly. Here, we propose to examine how these signaling pathways involving miR181a, AMPK and SIRTI regulate T effector and memory cell differentiation by controlling the activity of selected transcription factors and epigenetic imprinting. In four specific aims, we will perform mechanistic in vitro studies in parallel to correlative in vivo studies on antigen-specific T cells after yellow fever vaccination. In Aim 1, we will further define the role of AMPK and SIRTI pathways for T cell expansion and differentiation in young and elderly adults and examine the relationship to the expression of miR181a that declines with age. In Aim 2, we will explore how innate immunity and in particular type 1 IFN regulates these pathways and what the consequences are for the differentiation and survival of elderly CD4 T cells that have reduced sensitivity to respond to IFN. In Aim 3, we will determine the consequences of AMPK and SIRT pathway activation after TCR stimulation for chromatin remodeling and transcription factor binding. We will use a novel multidimensional epigenomic assay that relies on in vitro transposition of sequencing tags Into regions of accessible chromatin and that allows mapping the open chromatin and foot-printing transcription factor binding in antigen-specific T cells. In Aim 4, we will examine epigenetic signatures and miRNA profiles in CD28' effector T cells early after vaccination compare to CD28 T cells that arise with aging and chronic latent infections to explore whether these two populations are related or differ in the expression of genes involved in cell longevity and tissue residency.